Expansion and CD2/CD3/CD28 stimulation enhance Th2 cytokine secretion of human invariant NKT cells with retained anti-tumor cytotoxicity

Kelly Andrews, Anouk A.J. Hamers, Xiaodian Sun, Geoffrey Neale, Katherine Verbist, Paige Tedrick, Kim E. Nichols, Shalini Pereira, Daniel E. Geraghty, Asha B. Pillai

Research output: Contribution to journalArticlepeer-review

2 Scopus citations


Background aims: Key obstacles in human iNKT cell translational research and immunotherapy include the lack of robust protocols for dependable expansion of human iNKT cells and the paucity of data on phenotypes in post-expanded cells. Methods: We delineate expansion methods using interleukin (IL)-2, IL-7 and allogeneic feeder cells and anti-CD2/CD3/CD28 stimulation by which to dependably augment Th2 polarization and direct cytotoxicity of human peripheral blood CD3+Vα24+Vβ11+ iNKT cells. Results: Gene and protein expression profiling demonstrated augmented Th2 cytokine secretion (IL-4, IL-5, IL-13) in expanded iNKT cells stimulated with anti-CD2/CD3/CD28 antibodies. Cytotoxic effector molecules including granzyme B were increased in expanded iNKT cells after CD2/CD3/CD28 stimulation. Direct cytotoxicity assays using unstimulated expanded iNKT cell effectors revealed α-galactosyl ceramide (α-GalCer)-dependent killing of the T-ALL cell line Jurkat. Moreover, CD2/CD3/CD28 stimulation of expanded iNKT cells augmented their (α-GalCer-independent) killing of Jurkat cells. Co-culture of expanded iNKT cells with stimulated responder cells confirmed contact-dependent inhibition of activated CD4+ and CD8+ responder T cells. Discussion: These data establish a robust protocol to expand and novel pathways to enhance Th2 cytokine secretion and direct cytotoxicity in human iNKT cells, findings with direct implications for autoimmunity, vaccine augmentation and anti-infective immunity, cancer immunotherapy and transplantation.

Original languageEnglish (US)
Pages (from-to)276-290
Number of pages15
Issue number5
StatePublished - May 2020


  • NKT cells
  • co-stimulation
  • cytotoxicity
  • iNKT cells
  • immunotherapy
  • leukemia
  • regulatory T cells
  • transplantation

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology
  • Oncology
  • Genetics(clinical)
  • Cell Biology
  • Transplantation
  • Cancer Research


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